The present invention is directed to a polymeric composition that is capable of releasing nitric oxide (NO). The present invention is more particularly directed to a chitosan-based polymeric nitric oxide donor composition comprising a modified chitosan polymer and a nitric oxide [N2O2]xe2x88x92dimer, which provides site specific delivery and controlled release of nitric oxide under physiological conditions; pharmacological compositions comprising the polymeric composition; and the use of the polymeric composition to treat biological disorders for which an effective dosage of exogenous nitric oxide is indicated as a preferred method of treatment.
The chemical versatility of the nitric oxide (NO) molecule allows it to participate in a variety of physiologic processes. Nitric oxide (NO) reacts in biological systems with molecular oxygen (O2), superoxide (O2xe2x88x92) and transition metals to form high nitrogen oxides (NOx), peroxynitrite (OONOxe2x88x92) and metal-nitrosyl adducts which have various toxicities and biological activities.
The role of the nitric oxide (NO) molecule has been implicated in many physiologic processes, such as regulation of pulmonary hypertension via endothelium derived relaxing factor (EDRF)-induced vascular relaxation, central and peripheral neurotransmission, modulation of intestinal motility, regulation of male erectile function, macrophage-induced cytostasis and cytotoxicity, platelet inhibition, and non specific immune response.
Nitric oxide synthases (NOS) catalyze the oxidation of the substrate L-arginine into L-citrulline and nitric oxide (NO) in a variety of mammalian cell populations, such as pulmonary mucosa, submucosa, muscle, nerve and endothelium, reflecting the diverse biological activities of nitric oxide (NO).
Investigations have been conducted which pharmacologically manipulate the tissue level of nitric oxide (NO) by exogenous delivery of nitric oxide (NO) through inhalation of nitric oxide (NO) gas into the lungs via an endotracheal tube during mechanical ventilation. Although inhalation of nitric oxide gas has been useful in the treatment of such conditions as pulmonary hypertension, there are several disadvantages and limitations with this particular mode of therapy. Nitric oxide (NO) inhalation therapy requires large gas tanks, expensive monitoring equipment, and a highly trained and skilled technician to operate the tanks and equipment, to deliver a therapeutically effective amount of nitric oxide (NO) gas to a patient. Due to the volatility, hazardous and poisonous properties of nitric oxide (NO) gas, safe delivery of exogenous nitric oxide (NO) has been difficult to achieve.
In response to the great need in the art of nitric oxide (NO) therapy to develop a more convenient, less expensive and safer method to deliver exogenous nitric oxide (NO) to patients, numerous donor compounds have been developed to administer an effective dosage of exogenous nitric oxide (NO) in biological systems under physiological conditions.
A number of compounds have been developed which are capable of binding, delivering and releasing nitric oxide (NO) in physiological conditions upon being metabolized, including nitrovasodilators such as glyceryl trinitrite and sodium nitroprusside. (Ignarro et al., J. Pharmacol. Exp. Ther., 218, 739-749 (1981); Ignarro, Annu. Rev. Pharmacol. Toxicol., 30, 535-560 (1990); Kruszyna et al., Toxicol. Appl. Pharmacol., 91, 429-438 (1987); Wilcox et al., Chem. Res. Toxicol., 3, 71-76 (1990)). Although compounds such as glyceryl trinitrite and sodium nitroprusside are relatively stable under physiological conditions, a tolerance to glyceryl trinitrite via the exhaustion of the relevant enzyme/cofactor system is sometimes experienced in some applications, thus its use as a nitric oxide donor is limited. (Ignarro et al., Annu. Rev. Pharmacol. Toxicol., 25, 171-191 (1985); Kuhn et al., J. Cardiovasc. Pharmaol., 14 (Suppl. 11), S47-S54 (1989)). Furthermore, toxicity can develop from prolonged administration of sodium nitroprusside due to the metabolic production of cyanide. (Smith et al., xe2x80x9cA Potpourri of Biologically Reactive Intermediatesxe2x80x9d in Biological Reactive Intermediates IV. Molecular and Cellular Effects and Their Impact on Human Health (Witmer et al., eds.), Advances in Experimental Medicine and Biology Volume 283 (Plenum Press: New York, 1991), pp. 365-369).
A number of biodegradable compounds, such as primary, secondary, and polyamines, have been developed which are capable of delivering nitric oxide (NO), and releasing nitric oxide (NO) in physiologic conditions upon being metabolized. Keefer et al, U.S. Pat. No. 4,954,526 disclose a method of treating cardiovascular disorders with an effective amount of a stabilized complex formed from nitric oxide and primary amines and esters, ethers, of the formula [Rxe2x80x94N(H)N(NO)xe2x80x94]yX.
Further compounds have been developed that comprise diazenium diolates (NONOates) bound to polymers. NONOates are complexes of nitric oxide (NO) and nucleophiles (Xxe2x88x92) in which a nitric oxide (NO) dimer is bound to the nucleophilic residue via a nitrogen atom, thus forming a NONOate having the following chemical formula: 
Wherein R and Rxe2x80x2 are the same or different, and constitute any of a number of known organic moieties.
Traditionally, the synthesis of NONOates was accomplished by dissolving a nucleophile in an organic solvent and exposing the solution to a few atmospheres of NO gas for a period of a few days. A nucleophile, as the term is used above, is defined as an ion or molecule that donates a pair of electrons to an atomic nucleus to form a covalent bond. (Hawley""s Condensed Chemical Dictionary, Twelfth Edition). Useful nucleophiles for the synthesis of NONOates have traditionally included primary, secondary or polyamines. When a polyamine, such as spermine is used as the nucleophile, zwitterions are formed. Upon exposure of the nucleophile solution to NO gas, the NO dimer behaves as an electron pair acceptor, thus forming a covalent bond with the electron pair-donating nucleophile. This reaction results in a nitric oxide/nucleophile complex. The nucleophile moiety of the nitric oxide/nucleophile adduct can be further be bound to a polymer, such as a polysaccharide, such that the nucleophile moiety of the nitric oxide/nucleophile complex forms part of the polymer itself. NONOates are useful as nitric oxide donors in biological systems due to their ability to spontaneously disassociate under physiological conditions to regenerate the free nucleophile and molecular nitric oxide (NO).
Keefer et al, U.S. Pat. Nos. 5,250,550 and 5,155,137 disclose complexes of nitric oxide (NO) and polyamines, such as spermine and spermadine, useful in treating cardiovascular disorders, such as pulmonary hypertension, and which release nitric oxide (NO) under physiological conditions in a sustained and controlled manner. These complexes are made into pharmaceutical compositions by combination with medically acceptable carriers or diluents. Specifically, these compounds can be prepared into injectables by dissolving, suspending or emulsifying them in an aqueous or non-aqueous solvent.
Keefer et al, U.S. Pat. No. 5,212,204 discloses a method for lowering blood pressure using an antihypertensive composition comprising the N2O2xe2x88x92 functional group, an inorganic or organic moiety and a pharmaceutically acceptable cation. The organic or inorganic moiety is any moiety that will form the anithypertensive composition and release nitric oxide (NO) under physiological conditions upon decomposition.
Keefer et al, U.S. Pat. Nos. 5,208,233 and 5,039,705 disclose a method of treating cardiovascular disorders that will respond to a decrease in blood pressure, such as chronic hypertension, acute congestive heart failure, angina, acute myocardial infarction, left ventricular failure, cerebrovascular insufficiency and intracranial hemorrhage, by using an antihypertensive composition of secondary amines and nitric oxide adducts.
Keefer et al, U.S. Pat. No. 5,366,997 disclose a cardiovascularly active composition possessing antihypertensive properties comprising oxygen substituted derivatives of nucleophile-nitric oxide adducts as nitric oxide donor prodrugs.
Christodoulou et al, U.S. Pat. No. 5,389,675 disclose mixed ligand metal complexes of nitric oxide-nucleophile adducts which are capable of releasing nitric oxide (NO), and are useful as cardiovascular agents. The nitric oxide-nucleophile complex ligand are coordinated via the oxygen donor atoms of the bidentate N2O2xe2x88x92 functionality to metal centers, which are further bound to one or more additional ligands.
Smith et al, U.S. Pat. No. 5,691,423 disclose a polymeric composition capable of releasing nitric oxide (NO) in which the nitric oxide releasing functional group [N2O2]xe2x88x92 is bound to the polysaccharide via a nucleophile moiety or residue.
Many of the nitric oxide-nucleophile (NONOates) complexes have been promising as pharmacological compounds because, unlike, nitrovasodialators such as glyceryl trinitrite and sodium nitroprusside, they spontaneously release nitric oxide (NO) in aqueous solutions under physiological conditions without first having to be metabolized.
Although the NONOate complexes release molecular nitric oxide (NO) without first having to be metabolized, pharmacological applications have been limited by their propensity to distribute evenly throughout a given medium and their spontaneous release of nitric oxide (NO) in aqueous media, thus compromising site specific delivery of nitric oxide (NO) to target tissues.
Accordingly, there remains a great need to develop a low cost, readily biodegradable, biocompatible nitric oxide donor polymer composition comprising a nitric oxide [N2O2]xe2x88x92 dimer and a medically beneficial carrier molecule, capable of improved site specific delivery and controlled release of nitric oxide (NO) to target tissues under physiological conditions, without the further side effects of the nitric oxide donor compounds disclosed in the prior art.
It is an object of the present invention to provide a chitosan-based polymeric nitric oxide donor composition comprising a modified chitosan polymer and a nitric oxide [N2O2]xe2x88x92 dimer.
It is a further object of the present invention to provide chitosan-based polymeric nitric oxide donor compositions in which the nitric oxide [N2O2]xe2x88x92 dimer is bound directly to the modified chitosan polymer.
It is a further object of the present invention to provide a chitosan-based polymeric nitric oxide donor composition which further includes a medically beneficial carrier molecule.
It is a further object of the present invention to provide a chitosan-based polymeric nitric oxide donor composition capable of site specific delivery and controlled release of nitric oxide under physiological conditions.
It is a further object of the present invention to provide chitosan-based polymeric nitric oxide donor compositions having first order nitric oxide release kinetics.
It is a further object of the present invention to provide a method of preparing a chitosan-based polymeric nitric oxide donor composition comprising reacting a nitric oxide (NO) dimer with a modified chitosan polymer.
It is a further object of the present invention to provide a chitosan-based polymeric composition to treat respiratory distress, emphysema, external wounds, internal wounds and for coating vascular grafts.
These and other objects of the present invention, together with the advantages over the prior art relating to nitric oxide donor compositions, which should be become apparent from the specification which follows, are accomplished by the composition and method which is hereafter described and claimed.
The present invention, therefore, provides a chitosan-based polymeric nitric oxide donor composition comprising a modified chitosan polymer and a nitric oxide [N2O2]xe2x88x92 dimer, in which the nitric oxide [N2O2]xe2x88x92 dimer is bound directly to a nitrogen atom in the backbone of the modified chitosan polymer. The chitosan-based nitric oxide donor composition of the present invention further comprises a medically beneficial carrier molecule. The chitosan-based nitric oxide donor composition has first order nitric oxide release kinetics and provides site specific delivery and controlled release of nitric oxide under physiological conditions.
The nitric oxide donor composition of the present invention comprises a chitosan polymer and a nitric oxide [N2O2xe2x88x92] dimer, said composition having the general formula: 
wherein R1 is an organic group having from 1 to about 18 carbon atoms;
wherein x is from about 0.5 to about 0.8;
wherein y is from about 0.2 to about 0.5; and
wherein x and y are the mole fractions of each unit and the sum of x and y is 1.
The present invention further provides a method for preparing a nitric oxide donor composition comprising reacting a nitric oxide [N2O2]xe2x88x92 dimer with a chitosan polymer, said chitosan polymer having the following general formula: 
wherein R1 is an organic group having from 1 to about 18 carbon atoms;
wherein x is from about 0.5 to about 0.8;
wherein y is from about 0.2 to about 0.5; and
wherein x and y are the mole fractions of each unit and the sum of x and y is 1.
In one preferred embodiment, the present invention further provides a method for the preparing a nitric oxide donor composition comprising reacting nitric oxide (80-100 p.s.i.) with a chitosan polymer in the presence of sodium methoxide at room temperature, said chitosan polymer having the following general formula: 
wherein R1 is an organic group having from 1 to about 18 carbon atoms;
wherein x is from about 0.5 to about 0.8;
wherein y is from about 0.2 to about 0.5; and
wherein x and y are the mole fractions of each unit and the sum of x and y is 1.